NMR Verification of Structures of Bacterial Saccharide Precursors for Vaccines

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Three fractions of the O-specific polysaccharide(O-SP) isolated from Shigella sonnei have been investigated by NMR at 500 MHz, and the structure of the polysaccharide verified by 2D COSY, HSQC, and TOCSY methods, in conjunction with 1D 1H and 13C NMR, and 1H-coupled 13C NMR for confirmation of the anomeric configuration of the sugar residues. The 300 MHz 1H NMR spectra of about 40 batches of monosaccharide intermediates and linkers directed towards preparation of a conjugate vaccine for Vibrio cholera have been inspected for correct interpretation and authenticity. The degree of spectral dispersion (spreading of the NMR spectrum) and spectral resolution (narrowness of the resonances) is limited at 300 MHz, so the NMR data for the Vibrio cholera intermediates have been supplemented by 10 sets of 1H and/or 13C NMR data measured at 500 MHz. These spectra provide more detailed structural information and spectral assignments than are available from 300 MHz NMR spectra, for example, the detection of small, long-range 1H1H coupling constants, if present. The structures of five chromatographic fractions of Klebsiella pneumoniae type 2 capsular polysaccharide have been characterized by high-resolution NMR spectroscopy at 500 MHz, including the use of 1H NMR, 13C NMR, and 2D heteronuclear single quantum correlation (HSQC) NMR to correlate and assign the 1H and 13C chemical shifts. The NMR spectra provided information on the general types of structures involved and their purity, and afforded comparisons with literature data for related materials.